Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, United States.
ACS Chem Biol. 2011 Jan 21;6(1):47-60. doi: 10.1021/cb100358f. Epub 2010 Dec 28.
In the 12 years since the process of RNA interference (RNAi) was first discovered, great progress has been made in understanding its mechanism and exploiting its ability to silence gene expression to study gene function at a genome-wide level. Its extensive use as a screening method has yielded many published lists of genes that play novel roles in higher eukaryotes. However, the usefulness of this information is potentially limited by the occurrence of unintended off-target effects. Here we review the potential causes of off-target effects and the impact of this phenomenon in interpreting the results of high-throughput RNAi screens. In addition to targeting the intended gene product, artificial short interfering RNAs (siRNAs) can produce off-target effects by down-regulating the expression of multiple mRNAs through microRNA-like targeting of the 3' untranslated region. We examine why this phenomenon can produce high hit rates in siRNA screens and why independent validation of screening results is critical for the approach to yield new biological insights.
自 RNA 干扰 (RNAi) 过程首次被发现以来的 12 年中，人们在理解其机制以及利用其沉默基因表达的能力以在全基因组水平上研究基因功能方面取得了巨大进展。RNAi 已被广泛用作筛选方法，产生了许多已发表的基因列表，这些基因在高等真核生物中发挥着新的作用。然而，这种信息的有用性可能受到非预期的脱靶效应的限制。本文综述了脱靶效应的潜在原因，以及这种现象对解释高通量 RNAi 筛选结果的影响。除了靶向预期的基因产物外，人工短干扰 RNA (siRNA) 还可以通过 miRNA 样靶向 3'非翻译区来下调多个 mRNA 的表达，从而产生脱靶效应。我们研究了为什么这种现象会在 siRNA 筛选中产生高命中率，以及为什么对筛选结果进行独立验证对于该方法产生新的生物学见解至关重要。
